Synthetic duplex oligonucleotides, in particular short interfering RNAs (siRNAs), small hairpin RNAs (shRNAs), Piwi-interacting RNA (piRNA) mimics, and microRNA (miRNA) mimics, modulate gene expression via the RNA interference pathway (RNAi), leading to the cleavage of specific messenger RNAs (mRNAs). In the case of miRNA mimics, translational repression of specific mRNAs may take place in addition to, or instead of, cleavage of the mRNA.
Two factors that affect overall performance of synthetic duplex oligonucleotides are stability against nuclease degradation and the annealing strength. Chemical modifications have been identified that alter stability and annealing. Addition of O-alkyl (e.g., O-methyl) groups or halogens to the 2′ position of the ribose ring, and/or inter-nucleotide modifications to an oligonucleotide impedes degradation of these molecules by nucleases. At the same time, addition of 2′-O-alkyl groups can augment binding affinity and thus functionality.
Chemical modifications that can be added to the sense strand of an siRNA or miRNA mimic that enhance the stability and/or functionality of the duplex. See United States Patent Application Publication No. 2007/0269889, incorporated herein by reference in its entirety. These modifications, which comprise 2′-O-methyl modification of some or all of the nucleotides of the sense strand, minimize the nuclease sensitivity of the strand and enhance the entry of the antisense strand into the RNA interference silencing complex (RISC).
The inventors have now observed that addition of 2′-O-methyl modifications to the sense strand of e.g. miRNA mimics can, in some circumstances, have negative effects on that molecule. As these effects may be detrimental to the functionality of the molecule, it is desirable to identify modifications that can compensate for the negative properties.